Biodistribution of adeno‐associated virus type 2 carrying multi‐characteristic opsin in dogs following intravitreal injection

• Published in: Journal of cellular and molecular medicine Vol. 25; no. 18; pp. 8676 - 8686
• Main Authors: Tchedre, Kissaou T., Batabyal, Subrata, Galicia, Melissa, Narcisse, Darryl, Mustafi, Sourajit Mitra, Ayyagari, Ananta, Chavala, Sai, Mohanty, Samarendra K.
• Format: Journal Article
• Language: English
• Published: England John Wiley & Sons, Inc 01.09.2021; John Wiley and Sons Inc
• Subjects: Animals; Biodistribution; Body fluids; Clinical trials; Cloning; Deoxyribonucleic acid; Dependovirus - physiology; Disease; DNA; Dogs; Environmental restoration; Feces; Female; Gene therapy; Genetic Therapy - methods; Genetic Vectors; Genomes; Immune clearance; Immunohistochemistry; Inflammation; Injection; Lymph nodes; Male; MCO; Mutation; Original; Photoreceptors; Polymerase chain reaction; qPCR; rAAV; Retina; Retinal Diseases - therapy; retinal gene therapy; Saliva; Secretions; Spleen; Transplants & implants; Viruses; rAAV; MCO; biodistribution; qPCR; retinal gene therapy
• ISSN: 1582-1838; 1582-4934
• DOI: 10.1111/jcmm.16823

Gene therapy of retinal diseases using recombinant adeno‐associated virus (rAAV) vector‐based delivery has shown clinical success, and clinical trials based on rAAV‐based optogenetic therapies are currently in progress. Recently, we have developed multi‐characteristic opsin (MCO), which has been shown to effectively re‐photosensitize photoreceptor‐degenerated retina in mice leading to vision restoration at ambient light environment. Here, we report the biodistribution of the rAAV2 carried MCO (vMCO‐I) in live samples and post‐mortem organs following intraocular delivery in wild‐type dogs. Immunohistochemistry showed that the intravitreal injection of vMCO‐I resulted in gene transduction in the inner nuclear layer (INL) but did not induce detectable inflammatory or immune reaction in the dog retina. Vector DNA analysis of live body wastes and body fluids such as saliva and nasal secretions using quantitative polymerase chain reaction (qPCR) showed no correlative increase of vector copy in nasal secretions or saliva, minimal increase of vector copy in urine in the low‐dose group 13 weeks after injection and in the faeces of the high‐dose group at 3–13 weeks after injection suggesting clearance of the virus vector via urine and faeces. Further analysis of vector DNA extracted from faeces using PCR showed no transgene after 3 weeks post‐injection. Intravitreal injection of vMCO‐I resulted in few sporadic off‐target presences of the vector in the mesenteric lymph node, liver, spleen and testis. This study showed that intravitreal rAAV2‐based delivery of MCO‐I for retinal gene therapy is safe.